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Bioengineering Seminar Series: Justin Legleiter
Friday, October 7, 2011
11:00 a.m.
Room 1200 Jeong H. Kim Engineering Bldg.
For More Information:
Professor Silvia Muro
muro@umd.edu

The Role of Surfaces in the Formation of Disease-Related Protein Aggregates

Justin Legleiter
Assistant Professor
Department of Chemistry
West Virginia University

There are a large and diverse number of diseases that are commonly classified as conformational diseases. The common feature of these diseases is the rearrangement of a specific protein to a non-native conformation that promotes aggregation and deposition within tissues and/or cellular compartments. Such diseases include Alzheimer’s disease (AD), Huntington’s disease (HD), Parkinson’s disease (PD), amyloidoses, the prion encephalopathies, and many more. A common structural motif in the majority of these diseases is the emergence of extended, β-sheet rich, proteinaceous fibrillar aggregates that are commonly referred to as amyloids. These fibrillar species are comprised of intertwined protofibrillar filaments, which often have globular, soluble protein aggregate precursors, more commonly referred to as oligomers. We are using atomic force microscopy to study the aggregation of the β-amyloid (Aβ) peptide associated with AD and mutant huntingtin (htt) proteins associated with HD, with an interest in the potential role cellular and subcellular surfaces may play in aggregation.

We explored the effect of astrocyte secreted lipoprotein particles (ASLPs) containing different isoforms of apolipoprotein E (apoE), of which the apoE4 allele is a major risk factor for the development of AD, on the aggregation of Aβ in the presence of total brain lipid extract bilayers. The apoE4 allele was less effective in protecting lipid bilayers from disruption compared with apoE3. Size analysis of apoE-containing ASLPs and mechanical studies of bilayer properties revealed that apoE-containing ASLPs modulate the mechanical properties of bilayers by acquiring some bilayer components (most likely cholesterol and/or oxidatively damaged lipids). Measurement of bilayer mechanical properties was accomplished with scanning probe acceleration microscopy (SPAM). These measurements demonstrated that apoE4 was also less effective in modulating mechanical properties of bilayers in comparison with apoE3. This ability of apoE to alter the mechanical properties of lipid membranes may represent a potential mechanism for the suppression of Aβ induced bilayer disruption. Further studies to determine how point mutations in Aβ influence the aggregate morphology and formation kinetics of Aβ at solid/liquid interfaces may also be presented.

Huntington’s disease (HD) is caused by an expansion above 35–40 polyglutamine (polyQ) repeats in the huntingtin (htt) protein and results in accumulation of inclusion bodies that contain fibrillar deposits of mutant htt fragments. Intriguingly, polyQ length is directly proportional to the propensity for htt to form fibrils and to the severity of HD, and is inversely correlated to the age of onset. Although the structural basis for htt toxicity is unclear, the formation, abundance and/or persistence of toxic conformers that mediate neuronal dysfunction and degeneration in HD must also be polyQ length-dependent. Here we used atomic force microscopy (AFM) to show that mutant htt fragments and synthetic polyQ peptides form oligomers in a polyQ length-dependent manner. Time-lapse AFM shows oligomers form before fibrils, are transient in nature, and are occasionally direct precursors to fibrils. However, the vast majority of fibrils appear to form by monomer addition that coincides with the disappearance of oligomers. Thus, oligomers must undergo a major structural transition that precedes fibril formation. We have also explored the role polyQ context plays in the interaction of htt with lipid membranes.

About the Speaker
Justin attended Murray State University (Murray, KY), where he earned his bachelor’s degree in Chemistry in 2000. While at Murray, he worked in the lab of Professor Harry Fannin, where he used inductively coupled plasma spectroscopy to study bio-accumulation of metals in a fresh water protozoan, p. magnifica.

Justin performed his PhD work in Physical Chemistry at Carnegie Mellon University (Pittsburgh, PA) under the supervision of Dr. Tomasz Kowalewski. There, he studied the physicochemical properties of the self-assembly of the β-amyloid peptide and other relevant biological macromolecules associated with Alzheimer’s disease. He was also involved in developing atomic force microcopy (AFM) techniques in both air and fluids, especially the use of higher harmonic in processing signals and the use of tip acceleration to study local mechanical/chemical properties at the nanoscale. These studies led to the development of scanning probe acceleration microscopy (SPAM).

Justin then spent three years at the Gladstone Institute of Neurological Disease at the University of California, San Francisco as a postdoctoral fellow in the lab of Dr. Paul Muchowski. There, his main focus was the structural analysis of aggregates formed by mutant huntingtin fragments with expanded polyglutamine domains implicated in Huntington’s disease. His studies included the characterization of the aggregation of mutant huntingtin (htt) fragments with various polyQ lengths on chemically varied surfaces utilizing both ex situ and in situ AFM. He also worked to understand the potential of molecular chaperones, such as Hsp70 and Hsp40, and anti-htt antibodies to modulate the formation and stability of assemblies formed by mutant htt fragments.

In August of 2008, Justin moved to West Virginia University as an assistant professor in the C. Eugene Bennett Department of Chemistry.

This Event is For: Graduate • Faculty • Post-Docs

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